For immunoassays on infection disease-related test items, serum has been used as a sample to be tested. However, it takes about 30 minutes to separate serum from whole blood, including time for blood coagulation and time for subsequent centrifugation.
Typical examples of immunoassays include a radioimmunoassay (RIA), an enzyme immunoassay (EIA) a particle agglutination immunoassay and a counting immunoassay. However, RIA and EIA need B(Bound form)/F(Free form) separation after antigen-antibody reaction, and therefore, require time and labor before the results of the assays are obtained.
A particle agglutination immunoassay is advantageous in that it requires only the mixing of a sample to be tested with a suspension of insoluble carrier particles (e.g., latex) sensitized with an antibody or an antigen. Therefore it does not require the B/F separation and can be performed by simple operation.
Further, a counting immunoassay which is used serum or blood plasma and is known as an assay for measuring the degree of particle agglutination, antigen- or antibody-combined latex particles are reacted with an antibody or an antigen in a sample, so that they agglutinate according to the amount of the antigen or antibody. Agglutinated latex particles are distinguished by their size and counted. The count of unagglutinated latex particles is represented by M (monomer), the count of at least two agglutinated latex particles is represented by P (polymer), and the sum of M and P is represented by T (total). P/T is calculated as a degree of agglutination. If a calibration curve is determined beforehand by measuring the degree of agglutination of a known concentration of an antigen or antibody, the amount of the antigen or antibody in the sample can be found from the degree of agglutination of the sample (Sinkai Etsuro et al.: Principle of Measurement of PAMIA, Sysmex J. Vol. 20, No. 1, p.p. 77-78 (1997)). Since the agglutinated particles are directly counted, this assay is more accurate than the turbidimetric immunoassay which detects optical changes in the entire test sample.
In recent years, however, it has become necessary to judge rapidly whether or not a patient is infected with virus hepatitis, HIV or the like, for example, in the case of emergency operation. Accordingly, it is demanded that test time from collection of blood up to obtainment of test results be shortened. Also there is demanded a simple immunoassay that realizes the obtainment of highly accurate test results.
Taking the shortening of the test time into consideration, it is more desirable to use whole blood collected from a patient than to use serum, as a sample for immunoassays.
For example, Japanese Unexamined Patent Publication No. HEI 10(1998)-48214 proposes a whole blood assay using a conventional latex agglutination method. According to this assay, a whole blood sample is hemolyzed and the resulting sample is tested on its immune reaction by a latex turbidimetric immunoassay. This assay can provide test results simply and rapidly.
However, according to this assay, since the whole blood is hemolyzed, hemoglobin or fragments of hemolyzed blood cells interfere with the turbidimetric immunoassay using optical means and limit the accuracy of the assay. Therefore, the assay is not suitable for cases which require relatively high accuracy, for example, infection disease-related test items.
Whole blood usually contains about 40 to 50% of blood cell components in terms of hematocrit value. If whole blood is used in the same amount as in the case where serum is used, its measured values become lower than the values obtained by serum, reflecting the amount of blood cell components (a blood cell volume ratio content). In order to obtain measured values equivalent to those obtained by serum measurement, the values need to be corrected for the blood cell volume ratio content.
For this purpose, in Japanese Unexamined Patent Publication No. HEI 10 (1998)-48214, the hematocrit value is separately measured, and the measured values are corrected for the measured hematocrit value.
However, it is complicated to measure blood cells separately for correction. To avoid this, an inmmunoassay apparatus may be provided with a blood cell counting section and an immunoassay section, but such an apparatus becomes complicated and expensive. Therefore, there is a demand for a method for estimating or correcting influence by blood cells only by the immunoassay section.
Fully automated immunoassay apparatuses currently used are mostly designed for assaying serum (or blood plasma) samples, and therefore, do not have a stirrer which is necessary for assaying whole blood samples. If such apparatuses are used for immunoassays of whole blood samples, the blood cell components settle and only supernatant is used. The measured values are not affected seriously by the hematocrt value and may be not shifted apart to lower values.
In this case, if measured values are corrected for the hematocrit value separately measured by a blood cell counter for whole blood measurement as disclosed in Japanese Unexamined Patent Publication No. HEI 10 (1998)-48214, the measured values shift apart to higher values. As a result, precise measurement results cannot be obtained.
Alternatively, in an apparatus for both whole blood samples and serum samples which is already available commercially, a measuring sequence, an analysis program and the like are usually changed for measurement of a whole blood sample by designating the “whole blood measurement” on operation panel of the apparatus. However, where the number of samples is large, it is time- and labor-consuming to designate the “whole blood measurement” every time when a whole blood sample is used, and, where whole blood samples and serum samples are both used, it is also time- and labor-consuming to re-designate every time when the types of samples are changed. Furthermore, if the type of a sample is mis-designated, accurate measurement results are not obtained.